Over 30 million Americans suffer from moderate to profound sensorineural hearing loss (SNHL). SNHL is caused by a permanent degeneration of the sensory cells responsible for transmitting acoustic information to the brain: either the mechanosensory hair cells and/or the spiral ganglion (SG) neurons. There is no known cure for SNHL. Pallative treatments, such as hearing aids, are used to treat the symptoms of SNHL but no known treatment addresses the underlying cause of SNHL. Cochlear implants, which directly stimulate surviving SG neurons are an effective treatment for some portion of the population having SNHL. However, cochlear implantation is ineffective in nearly 15% of SNHL cases.
Pluripotent stem cells offer a promising approach to both model congenital deafness disorders and produce replacement inner ear sensory cells for curative therapy. Pluripotent cells, such as human embryonic stem (hES) cells and induced pluripotent stem (iPS) cells, can perpetually proliferate and differentiate into cells of each of the three embryonic germ layers (Thomson J, et al., “Embryonic stem cell lines derived from human blastocysts,” Science 282:1145-1147 (1998); Odorico J, et al., “Multilineage differentiation from human embryonic stem cell lines,” Stem Cells 19:193-204 (2001)). Differentiation of pluripotent cell cultures can occur spontaneously, which results in a seemingly random variety of cells (Watt F & Hogan Bm “Out of Eden: stem cells and their niches,” Science 287:1427-1430 (2000)). Alternatively, pluripotent cells can be induced to differentiate, e.g., by co-culturing the cells with cells of particular lineages or by chemical and/or mechanical detachment. The latter can be used to induce formation of embryoid bodies (EB), which, in turn, can differentiate into cells of multiple lineages.
Induced pluripotent (iPS) cells are generated by reprogramming somatic cells or differentiated progenitor cells to a state of pluripotency. Apart from their somatic cell origin, iPS cells share many characteristics of embryonic stem cells, such as the ability to grow perpetually and to differentiate into cells of the three germ layers. Like ES cells, iPS cells express one or more pluripotent cell-specific marker, such as OCT-4, SSEA-3, SSEA-4, TRA-1-60, TRA-1-81, and Nanog. iPS cells have been generated using retroviral vectors that randomly integrate into the target cell's DNA and using non-integrating vectors. iPS cells generated using non-integrating vectors are especially well suited for clinical application.
Generating inner ear cells from pluripotent cells is a significant challenge. Efforts to derive inner ear cells have been hampered by ill-defined culture systems that fail to recapitulate normal inner ear development mechanisms. During embryonic development the inner ear forms from a pool of progenitor cells called the preplacodal region. Previous studies aimed at deriving inner ear cells from pluripotent cells have failed to emphasize the importance of preplacodal cells, derived from non-neural ectoderm, in the differentiation path towards inner ear cells.
As can be appreciated from the above discussion, a need exists for methods to treat the cause of SNHL. Accordingly, it is one of the purposes of the present invention to replace degenerated sensory hair cells and/or auditory neurons, by providing a method for generating non-neural ectoderm, preplacodal ectoderm cells and otic placode cells.